TY - JOUR
T1 - The structure of the TNFRSF13C promoter enables differential expression of BAFF-R during B cell ontogeny and terminal differentiation
AU - Mihalcik, Stephen A.
AU - Huddleston, Paul M.
AU - Wu, Xiaosheng
AU - Jelinek, Diane F.
PY - 2010/7/15
Y1 - 2010/7/15
N2 - The B cell-activating factor of the TNF family receptor (BAFF-R), encoded by the TNFRSF13C gene, is critically important for transitional B cell survival to maturity. Thus, ligation of BAFF-R by BAFF delivers a potent survival signal. Reports implicating the BAFF/BAFF-R signaling axis in the pathogenesis of autoimmune human diseases and B lineage malignancies have largely prompted studies focusing on BAFF expression; however, there is an equally critical need to better understand BAFF-R expression. Initial BAFF-R expression, although characterized in murine B cells, has not yet been reported in human B lymphopoiesis. In this study, we first demonstrate that BAFF-R expression is absent from early precursors and is acquired by bone marrow B cells newly expressing the BCR. We next focused on identifying the specific genomic region that controls BAFF-R expression in mature B cells (i.e., the TNFRSF13C promoter). To accomplish this, we used in silico tools examining interspecies genomic conservation in conjunction with reporter constructs transfected into malignant B and plasma cell lines. DNase protection assays using nuclear extracts from BAFF-R - expressing cells suggested potential regulatory sites, which allowed the generation of EMSA probes that bound NFs specific to BAFF-R - expressing cells. With a more stringent analysis of interspecies homology, these assays identified a site at which a single nucleotide substitution could distinctly impact promoter activity. Finally, chromatin immunoprecipitation assays revealed the in vivo binding of the specific transcription factor c-Rel to the most proximal genomic region, and c-Rel small interfering RNA transfections in BAFF-R-expressing lines demonstrated a coincident knockdown of both c-Rel and BAFF-R mRNA.
AB - The B cell-activating factor of the TNF family receptor (BAFF-R), encoded by the TNFRSF13C gene, is critically important for transitional B cell survival to maturity. Thus, ligation of BAFF-R by BAFF delivers a potent survival signal. Reports implicating the BAFF/BAFF-R signaling axis in the pathogenesis of autoimmune human diseases and B lineage malignancies have largely prompted studies focusing on BAFF expression; however, there is an equally critical need to better understand BAFF-R expression. Initial BAFF-R expression, although characterized in murine B cells, has not yet been reported in human B lymphopoiesis. In this study, we first demonstrate that BAFF-R expression is absent from early precursors and is acquired by bone marrow B cells newly expressing the BCR. We next focused on identifying the specific genomic region that controls BAFF-R expression in mature B cells (i.e., the TNFRSF13C promoter). To accomplish this, we used in silico tools examining interspecies genomic conservation in conjunction with reporter constructs transfected into malignant B and plasma cell lines. DNase protection assays using nuclear extracts from BAFF-R - expressing cells suggested potential regulatory sites, which allowed the generation of EMSA probes that bound NFs specific to BAFF-R - expressing cells. With a more stringent analysis of interspecies homology, these assays identified a site at which a single nucleotide substitution could distinctly impact promoter activity. Finally, chromatin immunoprecipitation assays revealed the in vivo binding of the specific transcription factor c-Rel to the most proximal genomic region, and c-Rel small interfering RNA transfections in BAFF-R-expressing lines demonstrated a coincident knockdown of both c-Rel and BAFF-R mRNA.
UR - http://www.scopus.com/inward/record.url?scp=77955475763&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=77955475763&partnerID=8YFLogxK
U2 - 10.4049/jimmunol.1001120
DO - 10.4049/jimmunol.1001120
M3 - Article
C2 - 20554963
AN - SCOPUS:77955475763
SN - 0022-1767
VL - 185
SP - 1045
EP - 1054
JO - Journal of Immunology
JF - Journal of Immunology
IS - 2
ER -